Cell-restricted transcriptional modulators play critical roles in the process of selective gene regulation during hematopoiesis. We have been investigating the molecular and biological function of Erythroid Kr?ppel-like Factor (EKLF; KLF1). EKLF is a cell-restricted transcription factor that is essential for the erythroid program. Analysis of EKLF target gene regulation has revealed distinct transactivation mechanisms, and its protein interaction partners suggest additional unexplored roles in transcript control. The experiments of Aim 1 are directed at exploring the parameters of EKLF interactions with proteins and promoter DNA essential for these regulatory steps that lead to successful transcription. Analysis of EKLF's role in establishing an open chromatin structure, coupled with its interaction with histone H3 and the selective increase of H3.3 at the ?-globin promoter, suggest these observations are linked. As a result, the experiments of Aim 2 will address EKLF's role in coordinating histone H3.3 incorporation into chromatin. Analysis of the anemia in the heterozygous Nan mutant mouse has exposed an unexplained mechanism of gene-selective EKLF activation that leads to unique protein deficiency. The experiments of Aim 3 are designed to illuminate the molecular mechanism of the genetic distortion that results from the presence of Nan-EKLF in the erythroid cell. These studies will be aided by the use of in vivo assays, EKLF rescue systems, and primary or minimally manipulated cells. Elucidating EKLF's role in regulatory phenomena will continue to illuminate novel aspects of erythroid biology and the essential mechanisms by which a cell-restricted transcription factor can exert varied yet highly controlled influences on genetic expression and epigenetics.